C3B INACTIVATING POLYPEPTIDE
Polypeptides comprising a C3b binding region and a C3d inactivating region are disclosed, as well as nucleic acids and vectors encoding proteolytic such polypeptides. Also disclosed are cells and compositions comprising such polypeptides, and uses and methods using the same.
1 . A polypeptide comprising a C3b binding region and a C3b inactivating region.
2 . The polypeptide according to claim 1 , wherein the C3b inactivating region is capable of proteolytic cleavage of C3b.
3 . The polypeptide according to claim 1 or claim 2 , wherein the C3b inactivating region is capable of cleaving C3 α′ chain at positions 1303 and/or 1320.
4 . The polypeptide according to any one of claims 1 to 3 , wherein the C3b inactivating region comprises, or consists of, an amino acid sequence having at least 65% sequence identity to the amino acid sequence of SEQ ID NO:9.
5 . The polypeptide according to any one of claims 1 to 4 , wherein the C3b binding region binds to C3b in the region bound by a co-factor for Complement Factor I.
6 . The polypeptide according to any one of claims 1 to 5 , wherein the C3b binding region binds to C3b in the region bound by one of Complement Factor H, CR1, CD46, CD55 or C4-binding protein.
7 . The polypeptide according to any one of claims 1 to 6 , wherein the C3b binding region binds to C3b in the region bound by Complement Factor H, or the region bound by Complement Receptor 1 (CR1).
8 . The polypeptide according to any one of claims 1 to 7 , wherein the C3b binding region binds to C3b in the region bound by Complement Factor H complement control protein (CCP) domains 1-4, or the region bound by CR1 CCP domains 8-10 or 15-17.
9 . The polypeptide according to any one of claims 1 to 8 , wherein the C3b binding region comprises, or consists of, an amino acid sequence having at least 65% sequence identity to the amino acid sequence of SEQ ID NO:11, 13 or 14.
10 . The polypeptide according to any one of claims 1 to 9 , which is capable of diffusing across Bruch's membrane (BrM).
11 . The polypeptide according to any one of claims 1 to 10 , wherein the polypeptide is not glycosylated.
12 . The polypeptide according to any one of claims 1 to 11 , wherein the C3b inactivating region lacks an amino acid sequence conforming to the consensus sequence of SEQ ID NO:27.
13 . The polypeptide according to any one of claims 1 to 12 , wherein the polypeptide comprises a detection sequence, wherein the detection sequence comprises or consists of a cleavage site for a proteolytic enzyme, and wherein cleavage of the polypeptide with the proteolytic enzyme results in the production of a non-endogenous peptide.
14 . The polypeptide according to any one of claims 1 to 13 , comprising, or consisting of, an amino acid sequence having at least 65% sequence identity to the amino acid sequence of SEQ ID NO:32, 33, 34, 35, 36, 37, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 69, 70, 71, 72 or 73.
15 . The polypeptide according to any one of claims 1 to 14 , additionally comprising a secretory pathway sequence.
16 . The polypeptide according to claim 15 , wherein the secretory pathway sequence comprises one or more copies of an amino acid sequence conforming to the consensus sequence of SEQ ID NO:27, and wherein the polypeptide additionally comprises a cleavage site for removing the secretory pathway sequence.
17 . The polypeptide according to claim 16 , wherein the cleavage site for removing the secretory pathway sequence is a furin endoprotease cleavage site.
18 . A nucleic acid encoding the polypeptide according to any one of claims 1 to 17 .
19 . A vector comprising the nucleic acid of claim 18 .
20 . A cell comprising the polypeptide according to any one of claims 1 to 17 , the nucleic acid according to claim 18 , or the vector according to claim 19 .
21 . A method for producing a polypeptide, comprising introducing into a cell a nucleic acid according to claim 18 or a vector according to claim 19 , and culturing the cell under conditions suitable for expression of the polypeptide.
22 . A cell which is obtained or obtainable by the method according to claim 21 .
23 . A pharmaceutical composition comprising the polypeptide according to any one of claims 1 to 17 , the nucleic acid according to claim 18 , the vector according to claim 19 , or the cell according to claim 20 or claim 22 , and a pharmaceutically acceptable carrier, adjuvant, excipient, or diluent.
24 . The polypeptide according to any one of claims 1 to 16 , the nucleic acid according to claim 17 , the vector according to claim 18 , the cell according to claim 19 or claim 21 or the pharmaceutical composition according to claim 22 , for use in a method of treating or preventing a disease or condition.
25 . Use of the polypeptide according to any one of claims 1 to 17 , the nucleic acid according to claim 18 , the vector according to claim 19 , the cell according to claim 20 or claim 22 or the pharmaceutical composition according to claim 23 , in the manufacture of a medicament for treating or preventing a disease or condition.
26 . A method of treating or preventing a disease or condition, comprising administering to a subject the polypeptide according to any one of claims 1 to 17 , the nucleic acid according to claim 18 , the vector according to claim 19 , the cell according to claim 20 or claim 22 or the pharmaceutical composition according to claim 23 .
27 . A method of treating or preventing a disease or condition in a subject, comprising modifying at least one cell of the subject to express or comprise a nucleic acid according to claim 18 , or a vector according to claim 19 .
28 . The polypeptide, nucleic acid, vector, cell, or pharmaceutical composition for use according to claim 24 , the use according to claim 25 , or the method according to claim 26 to or claim 27 , wherein the disease or condition is a disease or condition in which C3b or a C3b-containing complex, an activity/response associated with C3b or a C3b-containing complex, or a product of an activity/response associated with C3b or a C3b-containing complex is pathologically implicated.
29 . The polypeptide, nucleic acid, vector, cell, or pharmaceutical composition for use, the use, or the method according to any one of claims 24 to 28 , wherein the disease or condition is age-related macular degeneration (AMD).
30 . A kit of parts comprising a predetermined quantity of the polypeptide according to any one of claims 1 to 17 , the nucleic acid according to claim 18 , the vector according to claim 19 , the cell according to claim 20 or claim 22 or the pharmaceutical composition according to claim 23 .
31 . A method of detecting a polypeptide in a sample, comprising:
(i) contacting a sample suspected to contain a polypeptide according to claim 13 with a proteolytic enzyme specific for the proteolytic cleavage site of the detection sequence; and
(ii) detecting the presence of the non-endogenous peptide.